tbuckley/cabot-search huggingface dataset:
OS10.7.A Activation of the CCR8-ACP5 axis by human microglia/macrophage derived CCL18 promotes glioma growth
Authors: Y Huang, E Motta, C Nanvuma, Y Yuan, L Kuhrt, P Xia, M Lubas, S Zhu, M Schnauss, F Hu, H Zhang, T Lei, M Synowitz, C Flüh, H Kettenmann
Journal: Neuro-Oncology (2022)

Abstract Background Glioblastoma multiforme is a highly malignant primary brain tumor with an average survival of 14 months and very limited therapeutic options. Glioma associated microglia and macrophages (GAMs) foster tumor growth by releasing several cytokines, which have only partly been identified. Here, we studied the chemokine (C-C motif) ligand 18 (CCL18), a chemokine which is only expressed in human, but not rodent GAMs, in a novel ex-vivo brain slice model including transplantation of human induced pluripotent stem cells (iPSC) derived human microglia (iMGL) and human glioma cells in to murine brain slices, which had been depleted of intrinsic murine microglia before. Material and Methods After establishing the humanized ex-vivo brain slice model, we performed immunohistochemical analysis (IHC) of growth and invasiveness, qrtPCR on glioma cells isolated by magnetic-activated cell sorting (MACS), functional assays measuring invasiveness, proliferation, migration and colony formation of glioma cells in vitro and in slice experiments. Corresponding studies on tumor growth and invasiveness were performed after treatment with a CCL18 neutralizing antibody, a CCR8 neutralizing antibodies and knockdown of CCR8, ACP5 (Acid Phosphatase 5) and PITPNM3 with small interfering RNA (siRNA) and short hairpin RNA (shRNA). QrtPCR, IHC and Westernblot analysis were performed on primary glioma specimens. We also conducted bioinformatic analyses, based on the TCGA GBM, GLIOVIS and GEPIA databases. Results We observed that CCL18 was highly expressed in GAMs, whereas CCR8 was only expressed in glioma cells. We identified the chemokine (C-C motif) receptor 8 (CCR8) as a functional receptor for CCL18 and ACP5 as an important down-stream signaling component in glioma cells. Activation of the CCL18/CCR8/ACP5 signaling pathway in human glioblastoma was associated with enhanced tumor growth and invasiveness. Conclusion GAMs derived CCL18 promoted glioma growth by activation of the CCR8/ACP5 axis in human glioma cells and therefore is a potential therapeutic target.
KS01.4.A Transcriptional evolution of glioblastoma point towards changes in bulk composition, mesenchymal sub-type as end-state, and a prognostic association with increased extracellular matrix gene expression
Authors: Y Hoogstrate, K Draaisma, L van Hijfte, S A Ghisai, I de Heer, I de Heer, M J van den Bent, P A Robe, P J French
Journal: Neuro-Oncology (2022)

Abstract Background Glioblastoma is the most prevalent and severe type of malignant brain tumor in adults. Although the genetic make-up initiating glioblastoma is increasingly better understood, a better understanding in the mechanisms that drive its evolution, heterogeneity and therapy resistance may reveal new directions for therapy development. To get better insights into glioblastoma evolution, we analyzed and deconvoluted transcriptomes of primary and recurrent glioblastoma resections. Material and Methods Matching primary and secondary resections from n=185 uniformly treated glioblastoma patients were collected as part of EORTC Study 1542 and RNA-sequenced. Data was extended with pairs from n=51 patients from the GLASS study. The datasets were subjected to differential and deconvolution analysis using in-house algorithms. Results When projecting the tumor samples into a reduced Glioblastoma Intrinsic Transcriptional Subtype space, visualization of transitions indicated that the CL subtype switches most often. As we found no more transitions from MES to other subtypes than to be expected by chance, we concluded that MES is an end-state. On average, tumor purity percentages decreased from ~67% to ~50%, mostly due to an increase in macrophages/microglia. Differential expression analysis was performed with correction for the fraction of non-malignant cells. While expression of glioblastoma associated oncogenes did not change significantly over time, marker genes for macrophages/microglia, neurons and oligodendrocytes were up-regulated whereas endothelial cell markers were down-regulated. A cluster of ~30 extracellular matrix associated (ECM) genes increased significantly over time. Single cell RNA-seq, IF-staining and RNA-ish indicated the signature is most strongly expressed near intra-tumoral vessels. Since endothelial marker genes were down-regulated over time, this suggests a form of prognostic vessel progression with a representative transcriptome signature. Conclusion Using a large cohort and validation set of uniformly treated patients, we demonstrate how the glioblastoma transcriptome changes over time with in particular changes the composition of the tumor and its environment. The tumor purity decrease over times suggests a more invasive phenotype or recruitment of non-malignant cells or a combination of both. A post-progression increase in expression of ECM-associated genes expressed near blood vessels was associated with poor survival. Concluding, while no consistent path for transcriptional evolution of tumor cells was observed other than transitions to the MES subtype, glioblastoma becomes more aggressive in time by (re-)organizing its environment.
P01.17.A Role of preoperative assessment in predicting tumor induced plasticity in patients with diffuse gliomas
Authors: F Latini, H Axelson, M Fahlström, M Jemstedt, Å Alberius Munkhammar, M Zetterling, M Ryttlefors
Journal: Neuro-Oncology (2022)

Abstract Background When diffuse gliomas (DG) affect the brain′s potential to reorganize functional networks, patients can exhibit seizures and/or language/cognitive impairment. The tumor-brain interaction and the individual connectomic organization cannot be predicted preoperatively. We aimed to, first, investigate the relationship between preoperative assessment and intraoperative findings of eloquent tumors in 36 DG operated with awake surgery. Second, we also studied possible mechanisms of tumor-induced brain reorganisation in these patients. Material and Methods FLAIR-MRI sequences were used for tumor volume segmentation and the Brain-Grid system (BG). This standardized radiological tool was used as an overlay for infiltration analysis. Neuropsychological (NPS) and/or language preoperative assessments were performed in all patients before the direct cortical/subcortical brain mapping. The distance between eloquent spots and tumor margins was registered and measured on the neuronavigation system. Tumours containing eloquent points within hyperintense area on FLAIR sequences (cortical or subcortical) were considered eloquent. Eloquent points within 5-10 mm from the FLAIR signal margin were considered peritumoral, while beyond 10 mm from the FLAIR signal were considered outside the tumour area. Pre and postoperative images (3 months after the operation) were merged to detect eventual residual tumour. Clinical, radiological, and topographical variables and intraoperative findings were used for correlation and logistic regression analyses. Results 84.6% of our patients displayed neuropsychological symptoms at the preoperative assessment. Eloquent tumors were detected in 75% of the cases. Impaired NPS functions were correlated with more invasive tumors, crucial location (A4C2S2/A3C2S2-voxels, left opercular-insular/sub-insular region) and higher risk of eloquent tumors. Epilepsy was correlated with larger tumor volumes and infiltrated A4C2S2 /A3C2S2 voxels. Language impairment was correlated with infiltrated A3C2S2 voxel. Group comparison between younger and older patients (cut-off at 38 years old based on ROC curves) showed a significative difference in the number of intratumoral cortical eloquent spots (p &amp;lt; .05) and the number of peritumoral cortical eloquent spots (p &amp;lt; .001). Conclusion Preoperative NPS impairment was linked with high risk of eloquent tumors. Epilepsy and language impairment were linked with specific tumor locations while peritumoral cortical eloquent spots reflected an early compensative mechanism with age as possible influencing factor. A systematic integration of extensive cognitive assessment and advanced neuroimaging can improve our comprehension of the connectomic brain organization at the individual scale and lead to a better oncological/functional balance.
OS10.6.A What is the initial cell in the subventricular zone for human glioblastoma genesis?
Authors: S Yoon, R Choi, S Cho, H Kim, J Lee, E Kim, J Chang, S Kim, H Kim, J Lee, S Kang
Journal: Neuro-Oncology (2022)

Abstract Background We all have a fundamental question about why glioblastoma (GBM) develops. In order to find the answer to this fundamental question, if you find out what the first cell is, you will get closer to the answer. GBM arises from the subventricular zone (SVZ). GBM is one of the most devastating tumour of human brain as the most optimal treatment barely prolongs the survival, and it does not cure the disease. As the majority of GBM tissues show copy number variations (CNV) of co-altered chromosomal 7 gain and 10 loss, we hypothesized the origin cell (Oc) of SVZ may be traced back with these markers. The cellular identity of the Oc is still unknown and it is different from the tumour-derived progenitor-like cells. We aimed to define these cells from the SVZ that have a potential to get activated into GBM. Material and Methods We compared bulk RNA sequencing (RNAseq) data of IDHwt GBM tumor tissue (n=122), tumor free SVZ from GBM patients (n=40), tumor-free control SVZ of non-glial tumor (n=9). Pared single nucleus RNAseq (snRNAseq) or single cell RNAseq (scRNAseq) samples of tumor free SVZ (n=11) and GBM tumor (n=8), were done to see cell specific CNVs. We developed genetically engineered mouse models for GBM genesis introducing three driver mutations (TP53, PTEN, and EGFRviii) into SVZ to isolate mouse Oc (mOc) and mouse cancer cells (mCc). The biological characteristics of separated mOc and mCc were compared. Bulk RNAseq and scRNAseq were performed on these cells (mOc, mCc), and their cellular state was compared with the human gene set. Results In this work, we found two types of the Oc in the RNA sequencing of 60 human tumour free-SVZ samples. Furthermore, single-cell level analysis revealed that two Oc types in SVZ harbor ongoing patterns of CNV co-alterations from Oc1 to Oc2, and finally to GBM. The Oc1 type cells contained the CNV signature of Oc2 ancestor with neural progenitor cell (NPC) signature. Oc2 type cells expressed a high level of EGFR than other cells with astrocyte-like cell signature. Both of these cells expressed oligodendrocyte progenitor cell (OPC)-like signatures in the SVZ. We validated the human-based findings by using the P53/PTEN/EGFR-mutant mouse model with EGFR/tdTomato overexpression and P53/PTEN knockout in the SVZ cells. As a result, non-tumourigenic and highly motile Oc-like cell-states are found in the mouse models, supporting the firework-like migration pattern from the SVZ. Conclusion Our results demonstrate how members of Oc preoccupy the SVZ, known as the stem cell niche and give rise to the tumour. We anticipate that a new therapy may emerge by targeting the Oc in the SVZ.
OS08.4.A Analysis of melanoma brain metastasis immune microenvironment through multiplex gene expression profiling
Authors: A A Ricci, G Collemi, G Orlando, C Tampieri, R Senetta, A Pellerino, F Bruno, A Melcarne, D Garbossa, R Rudà, R Soffietti, S Ribero, P Quaglino, P Cassoni, L Bertero
Journal: Neuro-Oncology (2022)

Abstract Background Novel immunotherapies based on targeting of specific immune checkpoints enabled a significant improvement of melanoma outcome, but melanoma brain metastases (BM) remain an unmet oncological need with an overall 2-year survival rate lower than 10%. Tumour immune microenvironment has been demonstrated to play a key role in BM establishment and development, but data regarding the specific milieu of melanoma BM is limited. Material and Methods Gene expression profiles of 55 samples of primary melanoma and BM were evaluated using the nCounter PanCancer IO 360 Panel (NanoString Technologies) targeting 770 mRNA involved in tumor immune microenvironment modulation. The case series consisted of 10 primary melanomas and their 10 matched BM, 25 unmatched BM, and 10 locally advanced control melanomas without evidence of BM after &amp;gt;5 year follow up. Results Among BM samples, most patients (25/45) were males and median age at BM diagnosis was 61,2 years with a median time to BM development of 2,1 years. Median OS from BM diagnosis was 1,3 years. Several genes resulted significantly downregulated in BM compared to primary melanomas, including SERPINB5 (p&amp;lt;0.001), ARG1 (p=0.0067), S100A8 (p&amp;lt;0.001), S100A9 (p&amp;lt;0.001), S100A12 (p=0.0037), IL1RN (p=0.0012), CCL21 (p=0.0012), CCL22 (p=0.0012) and CCL13 (p=0.037) and SELE (p=0.026); conversely, C7 was upregulated (p&amp;lt;0.001). Downregulated signatures in BM involved those associated with multiple immune cell populations, including neutrophils, dendritic cells, mast cells and Treg, as well as inflammatory chemokines, the CTLA4 immune checkpoint and ARG1 enzyme function; conversely, MAGEs-related signature was upregulated. Comparison between primary melanomas which developed BM and those which did not showed a significant overexpression of RRM2 (p=0.0247) and TNFRSF1A (p=0.032) genes in the latter group and an upregulation of the PD-1 pathway. Analysis according to tumour mutational status showed an upregulation of signatures associated with inflammatory chemokines, dendritic and myeloid cells and neutrophils. No differences were observed according to time to BM development and survival from BM diagnosis. Conclusion Our findings show that melanoma BM harbor distinct immunosuppressive mechanisms compared to primary tumors: this data elicits the importance of investigating the heterogeneity of BM microenvironment. Genes and pathways selectively overexpressed or downregulated in melanoma BM should be validated to be possibly considered as novel therapeutic targets.
P10.12.A CDK12/CDK13 inhibition disrupts a transcriptional program critical for glioblastoma survival
Authors: S Lier, I Rein, S Lund, A Lång, E Lång, N Meyer, A Dutta, S Anand, G Nesse, R Johansen, A Klungland, J Rinholm, S Bøe, A Anand, S Pollard, M Lerdrup, D Pandey
Journal: Neuro-Oncology (2022)

Abstract Background Glioblastoma multiforme (GBM) is the most prevalent and aggressive malignant tumor of the central nervous system. With a median survival of only one year, GBM patients have a particularly poor prognosis, highlighting a clear need for novel therapeutic strategies to target this disease. Transcriptional cyclin-dependent kinases (CDK), which phosphorylate key residues of RNA polymerase II (RNAPII) C-terminal domain (CTD), play a major role in sustaining aberrant transcriptional programs that are key to development and maintenance of cancer cells. Material and Methods We used pharmacological inhibition and genetic ablation to study effects of CDK12/CDK13 depletion on the proliferatory and migratory capacity of GBM cells and mouse xenografts. SLAM-seq, CUT&amp;RUN and cell cycle assays were used to study the mechanistic effects of CDK12/CDK13 depletion in GBM cells. Results CDK12/CDK13 depletion markedly reduced the proliferatory and migratory capacity of GBM cells, as well as in vivo growth. CDK12/CDK13 inhibition potentiated existing chemotherapeutic treatments. Mechanistically, inhibition of CDK12/CDK13 leads to a genome-wide abrogation of RNAPII CTD phosphorylation, which in turn disrupts transcription and cell cycle progression in GBM cells. Conclusion These results provide proof-of-concept for the potential of CDK12 and CDK13 as therapeutic targets for GBM.
P06.01.A Deconvolution of immunotherapy-treated glioblastoma identifies cellular heterogeneity and plasticity at the single-cell level
Authors: J D Hendriksen, A Flynn, S B Maarup, H S Poulsen, U Lassen, J Weischenfeldt
Journal: Neuro-Oncology (2022)

Abstract Background Glioblastoma is the most aggressive cancer originating in the brain with an average survival of 15 months. One of the characteristics of glioblastoma is the high level of intra-tumour heterogeneity (ITH), but the composition and complexity at the single-cell level is poorly understood. Here, we aimed to assess the effects and consequences of immune checkpoint inhibitor (ICI) on the cellular and molecular heterogeneity of glioblastoma tumours using at the single cell level. Material and Methods In collaboration with the phase I trials unit at Rigshospitalet, we performed paired molecular analysis of glioma cells from primary and relapse surgery after ICI treatment. Samples were analysed using single-cell RNA sequencing (scRNA-seq) as well as bulk RNA sequencing and whole exome DNA sequencing. Results In an effort to trace cellular lineages we developed and refined methods to a identify copy number changes using scRNA-seq. To this end, we identified clonal and subclonal tumour cell populations in each sample. We found high levels of ITH prior to treatment, both with respect to the glioblastoma subtype enrichment and the cell type-specific gene expression. Using expression-based cell-type classification, we found defined recurrent cell-type populations present at both surgery time points. The immune checkpoint treatment had consequences on the cellular phenotypes and proportions of tumour cells, suggesting a level of plasticity in the neoplastic cells. Moreover, we identified examples of clonal dynamics and sweeps following ICI treatment, pointing to potential treatment response and resistance in these population. Conclusion In summary, we pursued single cell-focused analysis of ICI treated glioblastoma patients to study the cellular and molecular heterogeneity within and between glioblastoma patients, which pointed to recurrent patterns of cellular responses following ICI treatment.
P07.04.B Heme biosynthesis factors and 5-ALA induced fluorescence: analysis of mRNA and protein expression in fluorescing and non-fluorescing gliomas
Authors: M Mischkulnig, T Roetzer-Pejrimovsky, D Lötsch-Gojo, N Kastner, K Bruckner, R Prihoda, A Lang, M Martinez-Moreno, J Furtner, A Berghoff, A Woehrer, W Berger, G Widhalm, B Kiesel
Journal: Neuro-Oncology (2022)

Abstract Background The intraoperative visualization of adult-type diffuse gliomas with 5-aminolevulinic acid (5-ALA) induced fluorescence is widely used in the neurosurgical field. While visible 5-ALA induced fluorescence is found in the majority of high-grade gliomas, most low-grade gliomas lack visible fluorescence during surgery. Recently, the heme biosynthesis pathway was identified as crucial influencing factor for presence of visible fluorescence since it metabolizes 5-ALA to fluorescing Protoporphyrin IX (PpIX). However, the exact alterations within the heme biosynthesis pathway resulting in visible 5-ALA induced fluorescence in gliomas are still unclear. The aim of the present study was thus to compare the mRNA and protein expression of promising intramitochondrial heme biosynthesis enzymes/transporters in glioma tissue samples of different fluorescence behavior. Material and Methods A total of 19 strongly fluorescing and 21 non-fluorescing tissue samples from neurosurgical adult-type diffuse gliomas (WHO grades II-IV) were included in the current analysis. In these samples, we investigated the mRNA expression by quantitative real time PCR and protein expression using immunohistochemistry of the intramitochondrial heme biosynthesis enzymes Coproporphyrinogen Oxidase (CPOX), Protoporphyrinogen Oxidase (PPOX), Ferrochelatase (FECH) and the transporter ATP-binding Cassette Subfamily B Member 2 (ABCG2). Results Regarding mRNA expression analysis, we found a significantly decreased ABCG2 expression in fluorescing specimens compared to non-fluorescing samples (p=0.001), whereas no difference in CPOX, PPOX and FECH was present. With respect to protein expression, significantly higher levels of CPOX (p=0.005), PPOX (p&amp;lt;0.01) and FECH (p=0.003) were detected in fluorescing samples. Similar to mRNA expression analysis, the protein expression of ABCG2 (p=0.001) was significantly lower in fluorescing samples. Conclusion Distinct alterations of the analyzed heme biosynthesis factors were found primarily on protein level. Our data indicate that heme biosynthesis pathway activity in general is enhanced in fluorescing gliomas with upregulation of PpIX generating enzymes and decreased ABCG2 mediated PpIX efflux outweighing the also increased further metabolization of PpIX to heme. Intramitochondrial heme biosynthesis factors thus constitute promising pharmacological targets to optimize intraoperative 5-ALA fluorescence visualization of usually non-fluorescing tumors such as low-grade gliomas.
P08.14.B Addressing and implementing coping strategies from pre-surgery can reduce anxiety and depression in the brain tumour population
Authors: v hurwitz, J p Lavrador, J La, L Brazil, O Al-Salihi, A Swampillai, R Bhangoo, F Vergani, K Ashkan, K Chia, S Hedges, A Suarez, E Kostick, C Robinson
Journal: Neuro-Oncology (2022)

Abstract Background Anxiety and depression is prevalent in oncology patients and disproportionately high in neuro-oncology. The screening tools are most frequently used when signs of depression are identified later in the patients pathway. The cognitive impact of a brain tumour means coping mechanisms are harder to integrate and retain. Anti-depressants and their impact on a brain tumour have long since been debated with no clear conclusion. Implementing a programme to address the anticipatory diagnosis of anxiety and depression from first contact potentially allows patients to discuss their mental health needs more openly. Material and Methods Patients are introduced to the concept of health related anxiety and depression following their first pre-assessment appointment, A combination of techniques are employed through different mediums including baseline assessment with EORTC QOL screening, MoCa assessment, CBT techniques, YouTube tutorials via the neuro-oncology cns channel and one to one sessions over six to nine sessions. Results Patients perceived quality of life is higher following completion of the programme. 75% of participants were able to identify coping strategies they had learnt and how and when to use them. Oncologists reported patients were better prepared for their results and initial oncology consenting appointment. High anxiety levels caused one participant to self-discharge prior to their surgery. On completion of four sessions they returned to the hospital and underwent awake craniotomy. Conclusion Identifying and implementing coping strategies at the earliest opportunity improves patients quality of life whilst living with a brain tumour. Anxiety and depression being discussed and the risk of developing either or both of these conditions allows for more effective prevention, awareness and diagnosis. There is the potential to further this work by looking at the impact on carers of early detection of patients anxiety and depression